We will study the kinetics of avian retrovirus RNA splicing in permissive chicken embryo fibroblasts and characterize in as much detail as possible the precursors, intermediates, and final products of this process. We will also determine the half-life of the viral mRNAs. Our approaches will be: 1) to carry out pulse-labeling and pulse-chase experiments with nucleoside precursors followed by the isolation of viral-specific RNA by hybridization to cloned viral DNA attached to cellulose. The viral specific RNA will then be analyzed by electrophoresis on agarose gel containing methyl mercury hydroxide; 2) to synthesize and process viral RNA in isolated nuclei from infected cells. We will try to determine the role of RNA methylations in the regulation of processing or the stability of avian retrovirus RNA. Our approaches will be to map the sites of the N6-methyladenosine residues on the genome RNA and on the viral mRNA, to investigate the effect of the in vivo methylation inhibitor cycloleucine on the kinetics of splicing and on mRNA stability, and to assay for the competence of undermethylated 38S genome RNA to rescue defective virions by microinjection into nuclei. We will try to determine whether the binding of p19 or pr76 to avian retrovirus RNA plays a role in the regulation of viral RNA processing. Our approach will be to test for the presence in infected cells of a complex of p19 or pr76 with pulse-labeled viral RNA. We will compare the translational efficiencies and do structural comparisons of the 38S RNA from virions, the intracellular 38S RNA that is associated with polysomes, and the 38S RNA that is not associated with polysomes.